Leucine Aminopeptidase Is Not Essential for Trimming Peptides in the Cytosol or Generating Epitopes for MHC Class I Antigen Presentation

Department of Pathology, University of Massachusetts Medical School, Worcester, MA 01655, USA.
The Journal of Immunology (Impact Factor: 4.92). 12/2005; 175(10):6605-14. DOI: 10.4049/jimmunol.175.10.6605
Source: PubMed


To detect viral infections and tumors, CD8+ T lymphocytes monitor cells for the presence of antigenic peptides bound to MHC class I molecules. The majority of MHC class I-presented peptides are generated from the cleavage of cellular and viral proteins by the ubiquitin-proteasome pathway. Many of the oligopeptides produced by this process are too long to stably bind to MHC class I molecules and require further trimming for presentation. Leucine aminopeptidase (LAP) is an IFN-inducible cytosolic aminopeptidase that can trim precursor peptides to mature epitopes and has been thought to play an important role in Ag presentation. To examine the role of LAP in generating MHC class I peptides in vivo, we generated LAP-deficient mice and LAP-deficient cell lines. These mutant mice and cells are viable and grow normally. The trimming of peptides in LAP-deficient cells is not reduced under basal conditions or after stimulation with IFN. Similarly, there is no reduction in presentation of peptides from precursor or full-length Ag constructs or in the overall supply of peptides from cellular proteins to MHC class I molecules even after stimulation with IFN. After viral infection, LAP-deficient mice generate normal CTL responses to seven epitopes from three different viruses. These data demonstrate that LAP is not an essential enzyme for generating most MHC class I-presented peptides and reveal redundancy in the function of cellular aminopeptidases.

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Available from: Margaret Karow, Jul 10, 2014
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    • "Trimming of the N-terminal leucine residue by ERAP1 was faster for larger peptides of >10 residues long, consistent with the length selection properties of ERAP1 (Fig. 3). In contrast, trimming by the cytosolic Leucine-aminopeptidase (LAP), a metabolic enzyme not essential for antigen generation (Lazaro et al., 2015; Towne et al., 2005), was independent of the length of the peptide, confirming the specialization of ERAP1 for larger peptides (Fig. 3). "
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    ABSTRACT: Endoplasmic Reticulum Aminopeptidase 1 (ERAP1) generates antigenic peptides for loading onto Major Histocompatibility Class I molecules (MHCI) and can regulate adaptive immune responses. During the last few years, many genetic studies have revealed strong associations between coding Single Nucleotide Polymorphisms (SNPs) in ERAP1 and common human diseases ranging from viral infections to cancer and autoimmunity. Functional studies have established that these SNPs affect enzyme activity resulting to changes in antigenic peptide processing, presentation by MHCI and cellular cytotoxic responses. These disease-associated polymorphisms are, however, located away from the enzyme's active site and are interspersed to different structural domains. As a result, the mechanism by which these SNPs can affect function remains largely elusive. ERAP1 utilizes a complex catalytic mechanism that involves a large conformational change between inactive and active forms and has the unique property to trim larger peptides more efficiently than smaller ones. We analyzed two of the most consistently discovered disease-associated polymorphisms, namely K528R and Q730E, for their effect on the ability of the enzyme to select substrates based on length and to undergo conformational changes. By utilizing enzymatic and computational analysis we propose that disease-associated SNPs can affect ERAP1 function by influencing: (i) substrate length selection and (ii) the conformational distribution of the protein ensemble. Our results provide novel insight on the mechanisms by which polymorphic variation distal from the active site of ERAP1 can translate to changes in function and contribute to immune system variability in humans. Copyright © 2015 Elsevier Ltd. All rights reserved.
    Molecular Immunology 07/2015; 67(2 Pt B). DOI:10.1016/j.molimm.2015.07.010 · 2.97 Impact Factor
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    • "Moreover, MHC class I levels at the cellmembrane were reduced after overexpression of TOP, there is more than just recycling beyond the proteasome and was increased upon inhibition of TOP with siRNA [51]. LAP is involved in N-terminal trimming of antigenic peptides [52] as it removes single amino acids from the N-terminus of larger peptides and can generate the model epitope SIINFEKL [52]. Although LAP can generate specific epitopes, LAP-deficient mice showed no differences in MHC class I levels when compared to wild-type mice [53], suggesting that LAP activity is not essential for antigen processing. "
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    ABSTRACT: Het proteasoom zorgt voor afbraak van eiwitten. Dat gebeurt in stappen – eerst genereert het proteasoom kleine peptiden die peptidases vervolgens verder afbreken tot individuele aminozuren. Een deel wordt gepresenteerd aan het immuunsysteem. Een subgroep daarvan is niet - zoals werd aangenomen - gemaakt door tripeptidyl peptidase II (TPPII) maar door metallopeptidases. Deze bevinding kan implicaties hebben voor het detecteren van kankercellen en cellen die door virussen geïnfecteerd zijn. Raspe onderzocht ook de functie van peptidases bij neurodegeneratieve ziekten als Alzheimer en Huntington die gepaard gaan met ophoping van moeilijk afbreekbare eiwitfragmenten. Bij Huntington bemoeilijkt een lange herhaling van hetzelfde aminozuur de afbraak van die eiwitfragmenten. Raspe vond twee peptidases (PSA en TPPII) die dergelijke ophopingen verminderen. PSA doet dat niet op de gebruikelijke wijze maar door het activeren van een ander opruimmechanisme, autofagie (waarbij de cel zichzelf als het ware opeet).
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    • "The peptides are then delivered to MHC I for antigen presentation to cytotoxic T-cells. While LAP clearly processes antigenic peptides in vitro (Beninga et al., 1998), analysis of LAP-knockout mice indicates that LAP activity is not essential for antigen presentation in vivo, suggesting that other aminopeptidases must be functionally redundant with LAPs (Towne et al., 2005). "
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    ABSTRACT: Leucine aminopeptidases (LAPs) are metallopeptidases that cleave N-terminal residues from proteins and peptides. While hydrolyzing Leu substrates, LAPs often have a broader specificity. LAPs are members of the M1 or M17 peptidase families, and therefore the LAP nomenclature is complex. LAPs are often viewed as cell maintenance enzymes with critical roles in turnover of peptides. In mammals, the M17 and M1 enzymes with LAP activity contribute to processing peptides for MHC I antigen presentation, processing of bioactive peptides (oxytocin, vasopressin, enkephalins), and vesicle trafficking to the plasma membrane. In microbes, the M17 LAPs have a role in proteolysis and have also acquired the ability to bind DNA. This property enables LAPs to serve as transcriptional repressors to control pyrimidine, alginate and cholera toxin biosynthesis, as well as mediate site-specific recombination events in plasmids and phages. In plants the roles of the M17 LAPs and the peptidases related to M1 LAPs are being elucidated. Roles in defense, membrane transport of auxin receptors, and meiosis have been implicated.
    Biological Chemistry 01/2007; 387(12):1535-44. DOI:10.1515/BC.2006.191 · 3.27 Impact Factor
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